Maximin-H5

Many pathogens, such as Pseudomonas aeruginosa and Escherichia coli bacteria can easily attach to surfaces and form stable biofilms. The formation of such biofilms in surfaces presents a problem in environmental, biomedical, and industrial processes, among many others. Aiming to provide a plausible solution to this issue, the anionic and hydrophobic peptide Maximin H5 C-terminally deaminated isoform (MH5C) has been modified with a cysteine in the C-terminal (MH5C-Cys) and coupled to polyethylene glycol (PEG) polymers of varying sizes (i.e., 2 kDa and 5 kDa) to serve as a surface protective coating. Briefly, the MH5C-Cys was bioconjugated to PEG and purified by size exclusion chromatography while the reaction was confirmed via SDS-PAGE and MALDI ToF. Moreover, the preventive antimicrobial activity of the MH5C-Cys-PEG conjugates was performed via the growth curves method, showing inhibition of bacterial growth after 24 h. The efficacy of these peptide-polymer conjugates was extensively characterized via scanning electron microscopy (SEM), minimum inhibition concentration (MIC), minimum biofilm inhibition concentration (MBIC), and minimum biofilm eradication concentration (MBEC) assays to evaluate their ability to eradicate and prevent the biofilms. Interestingly, this work demonstrated a critical PEG polymer weight of 5 kDa as ideal when coupled to the peptide to achieve inhibition and eradication of the biofilm formation in both bacteria strains. According to the MICs (40 μM) and MBICs (300 μM), we can conclude that this conjugate (MH5C-Cys-5 kDa) has an action that prevents/inhibits the formation of biofilms and the eradication of biofilms (MBEC 500 μM). In contrast, the MH5C-Cys peptide with PEG polymer of 2 kDa did not show inhibition or eradication of the biofilms.

Maximin H5 is an anionic antimicrobial peptide from amphibians, which carries a C-terminal amide moiety, and was found to be moderately haemolytic (20%). The α-helicity of the peptide was 42% in the presence of lipid mimics of erythrocyte membranes and was found able to penetrate (10.8 mN m(-1)) and lyse these model membranes (64 %). In contrast, the deaminated peptide exhibited lower levels of haemolysis (12%) and α-helicity (16%) along with a reduced ability to penetrate (7.8 m Nm(-1)) and lyse (55%) lipid mimics of erythrocyte membranes. Taken with molecular dynamic simulations and theoretical analysis, these data suggest that native maximin H5 primarily exerts its haemolytic action via the formation of an oblique orientated α-helical structure and tilted membrane insertion. However, the C-terminal deamination of maximin H5 induces a loss of tilted α-helical structure, which abolishes the ability of the peptide's N-terminal and C-terminal regions to H-bond and leads to a loss in haemolytic ability. Taken in combination, these observations strongly suggest that the C-terminal amide moiety carried by maximin H5 is required to stabilise the adoption of membrane interactive tilted structure by the peptide. Consistent with previous reports, these data show that the efficacy of interaction and specificity of maximin H5 for membranes can be attenuated by sequence modification and may assist in the development of variants of the peptide with the potential to serve as anti-infectives.

Here we report the first major example of anionic amphibian host defence peptides (HDPs) with anticancer activity. Maximin H5 (MH5N) is a C-terminally amidated, anionic host defence peptide from toads of the Bombina genus, which was shown to possess activity against the glioma cell line, T98G (EC50 = 125 μM). The peptide adopted high levels of α-helical structure (57.3%) in the presence of model cancer membranes (DMPC:DMPS in a molar ratio of 10:1). MH5N also showed a strong ability to penetrate these model membranes (Π = 10.5 mN m-1), which correlated with levels of DMPS (R2 > 0.98). Taken with the high ability of the peptide to lyse these membranes (65.7%), it is proposed that maximin H5 kills cancer cells via membranolytic mechanisms that are promoted by anionic lipid. It was also found that C-terminally deaminated maximin H5 (MH5C) exhibited lower levels of α-helical structure in the presence of cancer membrane mimics (44.8%) along with a reduced ability to penetrate these membranes (Π = 8.1 mN m-1) and induce their lysis (56.6%). These data suggested that the two terminal amide groups of native maximin H5 are required for its optimal membranolytic and anticancer activity.